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- EMDB-11971: Stressosome complex from Listeria innocua -

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Basic information

Entry
Database: EMDB / ID: EMD-11971
TitleStressosome complex from Listeria innocua
Map data
Sample
  • Complex: stressosome complex RsbR and RsbS
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbS protein
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbR protein
Function / homology
Function and homology information


oxygen binding / heme binding
Similarity search - Function
RsbS co-antagonist protein RsbRA N-terminal domain / Rsbr N terminal / STAS domain / STAS domain profile. / STAS domain / STAS domain superfamily / Globin/Protoglobin
Similarity search - Domain/homology
RsbS protein / RsbR protein
Similarity search - Component
Biological speciesListeria innocua Clip11262 (bacteria) / Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.86 Å
AuthorsMiksys A / Fu L / Madej MG / Ziegler C
Funding support Germany, 1 items
OrganizationGrant numberCountry
H2020 Marie Curie Actions of the European Commission721456 Germany
CitationJournal: Commun Biol / Year: 2022
Title: Molecular insights into intra-complex signal transmission during stressosome activation.
Authors: Algirdas Miksys / Lifei Fu / M Gregor Madej / Duarte N Guerreiro / Susann Kaltwasser / Maria Conway / Sema Ejder / Astrid Bruckmann / Jon Marles-Wright / Richard J Lewis / Conor O'Byrne / ...Authors: Algirdas Miksys / Lifei Fu / M Gregor Madej / Duarte N Guerreiro / Susann Kaltwasser / Maria Conway / Sema Ejder / Astrid Bruckmann / Jon Marles-Wright / Richard J Lewis / Conor O'Byrne / Jan Pané-Farré / Christine Ziegler /
Abstract: The stressosome is a pseudo-icosahedral megadalton bacterial stress-sensing protein complex consisting of several copies of two STAS-domain proteins, RsbR and RsbS, and the kinase RsbT. Upon ...The stressosome is a pseudo-icosahedral megadalton bacterial stress-sensing protein complex consisting of several copies of two STAS-domain proteins, RsbR and RsbS, and the kinase RsbT. Upon perception of environmental stress multiple copies of RsbT are released from the surface of the stressosome. Free RsbT activates downstream proteins to elicit a global cellular response, such as the activation of the general stress response in Gram-positive bacteria. The molecular events triggering RsbT release from the stressosome surface remain poorly understood. Here we present the map of Listeria innocua RsbR1/RsbS complex at resolutions of 3.45 Å for the STAS domain core in icosahedral symmetry and of 3.87 Å for the STAS domain and N-terminal sensors in D2 symmetry, respectively. The structure reveals a conformational change in the STAS domain linked to phosphorylation in RsbR. Docking studies indicate that allosteric RsbT binding to the conformationally flexible N-terminal sensor domain of RsbR affects the affinity of RsbS towards RsbT. Our results bring to focus the molecular events within the stressosome complex and further our understanding of this ubiquitous signaling hub.
History
DepositionNov 22, 2020-
Header (metadata) releaseDec 1, 2021-
Map releaseDec 1, 2021-
UpdateJul 20, 2022-
Current statusJul 20, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.735
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.735
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7b0u
  • Surface level: 0.6
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7b0u
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_11971.png

Downloads & links

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Map

FileDownload / File: emd_11971.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.0635 Å
Density
Contour LevelBy AUTHOR: 0.735 / Movie #1: 0.735
Minimum - Maximum-3.3991375 - 6.5165963
Average (Standard dev.)0.0034875653 (±0.24009643)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 408.38403 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06351.06351.0635
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z408.384408.384408.384
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ384384384
MAP C/R/S213
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-3.3996.5170.003

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Supplemental data

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Sample components

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Entire : stressosome complex RsbR and RsbS

EntireName: stressosome complex RsbR and RsbS
Components
  • Complex: stressosome complex RsbR and RsbS
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbS protein
    • Protein or peptide: RsbR protein
    • Protein or peptide: RsbR protein

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Supramolecule #1: stressosome complex RsbR and RsbS

SupramoleculeName: stressosome complex RsbR and RsbS / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Listeria innocua Clip11262 (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
Molecular weightTheoretical: 1.5 MDa

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Macromolecule #1: RsbR protein

MacromoleculeName: RsbR protein / type: protein_or_peptide / ID: 1 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Strain: ATCC BAA-680 / CLIP 11262
Molecular weightTheoretical: 31.704586 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP ...String:
MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP LIGTIDTERA KLIIENLLIG VVKNRSEVVL IDITGVPVVD TMVAHHIIQA SEAVRLVGCQ AMLVGIRPEI AQ TIVNLGI ELDQIITTNT MKKGMERALA LTNREIVEKE G

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Macromolecule #2: RsbR protein

MacromoleculeName: RsbR protein / type: protein_or_peptide / ID: 2 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Strain: ATCC BAA-680 / CLIP 11262
Molecular weightTheoretical: 31.784566 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP ...String:
MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP LIGTIDTERA KLIIENLLIG VVKNRSEVVL IDITGVPVVD TMVAHHIIQA SEAVRLVGCQ AMLVGIRPEI AQ (TPO)IVNLGI ELDQIITTNT MKKGMERALA LTNREIVEKE G

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Macromolecule #3: RsbS protein

MacromoleculeName: RsbS protein / type: protein_or_peptide / ID: 3 / Number of copies: 20 / Enantiomer: LEVO
Source (natural)Organism: Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Strain: ATCC BAA-680 / CLIP 11262
Molecular weightTheoretical: 12.606707 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString:
MGIPILKLGE CLLISIQSEL DDHTAVEFQE DLLAKIHETS ARGVVIDITS IDFIDSFIAK ILGDVVSMSK LMGAKVVVTG IQPAVAITL IELGITFSGV LSAMDLESGL EKLKQELGE

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Macromolecule #4: RsbR protein

MacromoleculeName: RsbR protein / type: protein_or_peptide / ID: 4 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Strain: ATCC BAA-680 / CLIP 11262
Molecular weightTheoretical: 31.864547 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP ...String:
MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP LIGTID(TPO)ERA KLIIENLLIG VVKNRSEVVL IDITGVPVVD TMVAHHIIQA SEAVRLVGCQ AMLVGIRP E IAQ(TPO)IVNLGI ELDQIITTNT MKKGMERALA LTNREIVEKE G

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Macromolecule #5: RsbR protein

MacromoleculeName: RsbR protein / type: protein_or_peptide / ID: 5 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Listeria innocua serovar 6a (strain ATCC BAA-680 / CLIP 11262) (bacteria)
Strain: ATCC BAA-680 / CLIP 11262
Molecular weightTheoretical: 31.784566 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP ...String:
MYKDFANFIR TNKKDLLNNW MNEMEKQSDP LINDIAKEPM YEETSIEFVD LIVSNITENG SKFNEKLDDF AEKVVHLGWP IHFVTTGLR VFGLLVYTAM RDEDLFLKRE EKPEDDAYYR FETWLSSMYN KVVTAYADTW EKTVSIQKSA LQELSAPLLP I FEKISVMP LIGTID(TPO)ERA KLIIENLLIG VVKNRSEVVL IDITGVPVVD TMVAHHIIQA SEAVRLVGCQ AMLVGIRP E IAQTIVNLGI ELDQIITTNT MKKGMERALA LTNREIVEKE G

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.6 mg/mL
BufferpH: 8.5
Component:
ConcentrationFormulaName
100.0 mMNaClsodium chloride
50.0 mMC4H11NO3tris

Details: pH 8.5
GridModel: Quantifoil / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Pressure: 0.029 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: INTEGRATING / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 3.86 Å / Resolution method: FSC 0.143 CUT-OFF / Software: (Name: RELION (ver. 3.0), cryoSPARC (ver. V2)) / Number images used: 32031
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Target criteria: Correlation coefficient
Output model

PDB-7b0u:
Stressosome complex from Listeria innocua

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